Die Hämoglobine von Myxine glutinosa L. (Cyklostomata) : Primärstruktur des monomeren Hämoglobins III und Untersuchung von Funktion und Evolution

Sammanfattning: Studies on the hemoproteins of the hagfish Myxine glutinosa L. (Cyclostomata) are of great interest with respect to the evolution of vertebrate hemoglobins.Hagfish hemoglobin is heterogeneous. Three main components, Hb I, Hb II and Hb III, as well as ten minor ones, have been isolated by isoelectric focusing. All main components are monomers, their molecular weights decreasing from Hb I to Hb III. Two of them, Hb I and Hb III, have been crystallized.The amino acid composition of the main components have been determined. They differ considerably from each other and from the hemoglobins of the lampreys, members of another subclass of Cyclostomata. The N-terminal residue of Hb III is proline, whereas those of Hb I and Hb II are blocked.The amino acid sequence of Hb III from Myxine has been elucidated. The globin was subjected to tryptic digestion, as well as to cleavage with cyanogen bromide and BNPS-skatol. The sequences of the separated peptides were determined by automated Edman degradation. Globin Hb III consists of 148 amino acid residues and contains no cysteine, in contrast to other vertebrate hemoglobins.A novel heme linkage has been demonstrated, which has only been found in opossum hemoglobin and globin from the invertebrate Chironomus. The regularly occurring distal histidine E7 is substituted by glutamine, and valine E11 by isoleucine. It has been proposed that Gin E7 is directed towards the outside of the heme pocket and Ile E11 towards the inside, replacing histidine in the distal heme contact.Globin Hb III has an additional segment of nine amino acid residues at the N-terminus in contrast to mammalian hemoglobins. This N-terminal nona- peptide is also present in globins from other Cyclostomata.The oxygen binding characteristics of Myxine hemoglobin have been studied. The main components show no cooperative oxygen binding and do not exhibit any Bohr effect, having however, different oxygen affinities. Although all hemoglobins of Cyclostomata are monomers, their oxygen binding characteristics vary. ESR studies of the nitrosyl complex of hemoglobin Hb III have shown that the critical substitutions in the heme linkage at E7 and E11 do not disrupt the proximal histidine-iron bond, even though substitutions of the same type are known to cause destabilization of the R quaternary structure in tetrameric mammalian nitrosylhemoglobins.The primary structure of Hb III differs by more than 50% from that of hitherto analysed lamprey hemoglobins. More than 400 million years were required for the development of this difference in the course of evolution. Nevertheless, a comparison of the hemoglobin sequences from the two branches of Agnatha, hagfish and lamprey, suggests a monophyletic origin, distinct from that of Gnathostomata. The primary structure of Myxine hemoglobin is compatible with the theory that this hemoglobin is a link between invertebrate and vertebrate globins, and that the divergence of hemoglobins occurred more than 500 million years ago.